The major histocompatability (MHC) class II molecules are expressed on the surfaces of antigen-presenting cells and B lymphocytes. By binding antigens and presenting the antigens to T cells, the MHC class II molecules are involved in triggering an immune response. Thus, the level of expression of the MHC class II molecules affects the induction of an immune response.
The genes which encode the xcex1 and xcex2 polypeptides of the MHC class II antigens are located at the HLA-D (histocompatability leukocyte antigen-D) region of the chromosome. Isotypes of the class II genes include those which are designated HLA-DR, HLA-DQ, and HLA-DP. Within these genes, there is substantial allelic polymorphism; for example, HLA-DR alleles include DR1, DR2, DR3, and DR4. In addition, subtypes of these alleles exist; for example, subtypes of DR4 include Dw4, Dw10, Dw13, Dw14, and Dw15.
Several autoimmune diseases are associated with expression of particular alleles of the MHC class II genes. For example, approximately 93% of patients afflicted with rheumatoid arthritis express HLA-DR1, HLA-DR4, or both (McDermott et al., Bulletin on the Rheumatic Diseases, 38:1-10). Other autoimmune diseases also are linked to expression of particular alleles. For example, Felty""s syndrome, Sjogren""s syndrome, systemic lupus erythematosus, and the development of toxicities to gold and penicillamine are associated with various HLA-DR alleles (McDermott et al., Bulletin on the Rheumatic Diseases, 38:1-10). As another example, pauciarticular juvenile rheumatoid arthritis is associated with HLA-PB2.1 (Begovich et al., 1989, PNAS 86:9489-9493). Approximately 70% of patients with insulin-dependent diabetes mellitus express HLA-DQ3.2B, DQA1, or DQB1, and susceptibility to the autoimmune dermatologic disease pemphigus vulgaris is linked to expression of HLA-DQB1.3 (Scharf et al., 1989, PNAS 86:6215-6219).
Activation of transcription of the MHC Class II genes has been shown to be dependent upon the transactivator CIITA (Steimle et al., 1993, Cell 75:135-146). CIITA does not function by directly binding DNA itself but rather by interacting with a conserved set of DNA binding proteins that associate with the class II promoter region (Steimle et al. (1994) Science 265:106-109; Chang et al. (1996) Immunity 4:167-178; Mach et al. (1996) Annu. Rev. Immuno. 14:301-331; Riley et al. (1995) Immunity 2:533-543). The transcriptional activation function of CIITA has been mapped to an amino terminal acidic domain (amino acids 26-137) (Zhou and Glimcher (1995) Immunity 2:545-553). The defect in a subset of MHC Class II deficient patients has been shown to be a mutation in CIITA, thereby demonstrating the importance of CIITA in regulating MHC Class II gene transcription (Steimle et al., 1993, Cell 75:135-146; Bontron et al. (1997) Hum. Genet. 99:541-546; Steimle et al. (1996) Adv. Immunol. 61:327-340). Given the critical role that CIITA plays in regulating MHC Class II gene expression, further information on how CIITA functions is of great interest.
This invention pertains to a protein that interacts with CIITA and enhances CIITA-regulated transciption from MHC class II gene promoters. A nucleic acid molecule encoding a protein that interacts with CIITA, termed CIITA-interacting protein 104 (also referred to herein as CIP104), has now been isolated and characterized. The CIP104-encoding nucleic acid was isolated based upon the ability of the encoded protein to interact with CIITA in a yeast two hybrid assay system. The nucleotide sequence of a cDNA encoding CIP104 and the predicted amino acid sequence of CIP104 have been determined and are shown in SEQ ID NOs: 1 and 2, respectively. Furthermore, CIP104 has been shown to enhance transcription from MHC class II promoters. This invention provides isolated compositions of CIP104 and isolated nucleic acid sequences encoding CIP104, as well as other compositions related thereto and methods of use thereof.
One aspect of the invention pertains to isolated nucleic acid molecules encoding CIP104, or fragments thereof. In one embodiment, the invention provides an isolated nucleic acid molecule comprising a nucleotide sequence encoding CIP104. In another embodiment, the invention provides an isolated nucleic acid molecule comprising a nucleotide sequence encoding a protein, wherein the protein comprises an amino acid sequence that is homologous to the amino acid sequence of SEQ ID NO: 2 and interacts with CIITA. In yet another embodiment, the invention provides an isolated nucleic acid molecule which hybridizes under stringent conditions to a nucleic acid molecule comprising the nucleotide sequence of SEQ ID NO: 1. In yet another embodiment, the invention provides an isolated nucleic acid molecule comprising the nucleotide sequence of SEQ ID NO: 1. In still other embodiments, the invention provides an isolated nucleic acid molecule encoding the amino acid sequence of SEQ ID NO: 2. Isolated nucleic acid molecules encoding CIP104 fusion proteins and isolated antisense nucleic acid molecules are also encompassed by the invention.
Another aspect of the invention pertains to vectors, such as recombinant expression vectors, containing an nucleic acid molecule of the invention and host cells into which such vectors have been introduced. In one embodiment, such a host cell is used to produce CIP104 by culturing the host cell in a suitable medium. If desired, CIP104 can be then isolated from the host cell or the medium.
Still another aspect of the invention pertains to isolated CIITA-interacting proteins, or portions thereof. In one embodiment, the invention provides an isolated CIP104 protein, or a portion thereof that interacts with CIITA. In yet another embodiment, the invention provides an isolated protein which comprises an amino acid sequence homologous to the amino acid sequence of SEQ ID NO: 2 and that interacts with CIITA. CIP104 fusion proteins are also encompassed by the invention.
The CIP104 proteins of the invention, or fragments thereof, can be used to prepare anti-CIP104 antibodies. Accordingly, the invention further provides antibodies that specifically binds CIP104. In one embodiment, the antibodies are polyclonal. In another embodiment, the antibodies are monoclonal. In yet another embodiment, the antibodies are labeled with a detectable substance.
The CIP104-encoding nucleic acid molecules of the invention can be used to prepare nonhuman transgenic animals that contain cells carrying a transgene encoding CIP104 or a portion of CIP104. Accordingly, such transgenic animals are also provided by the invention. In one embodiment, the CIP104 transgene carried by the transgenic animal alters an endogenous gene encoding endogenous CIP104 (e.g., a homologous recombinant animal).
Another aspect of the invention pertains to methods for detecting the presence of CIP104 activity (e.g., CIP104 protein or mRNA) in a biological sample. To detect CIP104 activity (e.g., protein or mRNA), the biological sample is contacted with an agent capable of detecting CIP104 protein (such as a labeled anti-CIP104 antibody) or CIP104 mRNA (such as a labeled nucleic acid probe capable of hybridizing to CIP104 mRNA) such that the presence of CIP104 protein or mRNA is detected in the biological sample.
Still another aspect of the invention pertains to methods for identifying compounds that modulate the activity or expression of CIP104 and methods for identifying compounds that modulate an interaction between CIP104 and CIITA. Screening methods for identifying proteins that interact with CIP are also encompassed by the invention.